This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. DESCRIPTION Mesial temporal lobe epilepsy (TLE) is the most common type of epilepsy in adults and can be caused by a variety of insults. Specific loss of entorhinal cortex (EC) Layer 3 (L3) pyramidal neurons (PNs), and damage elsewhere, as in the hippocampus, are characteristic for late stages of the disease. This project studies the cellular and synaptic changes that occur in the entorhinal cortex in the early stages of TLE evolution, poorly understood at present. The research utilizes the lithium-pilocarpine model of TLE to examine early mechanisms of EC L5 hyperexcitability. In this model, status epilepticus (SE) is evoked by systemic application of the muscarinic agonist pilocarpine and terminated after one hour by benzodiazepines. After a silent period of 2 [unreadable]4 weeks spontaneous seizures occur. Resulting seizures (both pilocarpine-induced and spontaneous) originate in EC-L5, and then spread to L2 and on into the hippocampus. Pathologic release of endogenous acetylcholine may also initiate similar status epilepticus, as a consequence of the dense cholinergic innervation of all layers of the entorhinal cortex by convergence of cholinergic fibers from the basal ganglia, the forebrain nuclei and the septum. Studies in this project test the hypothesis that deficiencies in synaptic inhibition and potentiation of synaptic excitation of EC-L5 pyramidal neurons cause TLE. Most studies compare preparations from control and pilocarpine treated rats 2 and 3 weeks after SE, to determine a) changes in neuronal Cl--transporters (compromising GABAergic synaptic inhibition), b) vesicular release probability changes for glutamatergic excitatory and GABAergic inhibitory synaptic terminals, and c) excitability changes in EC L5 neuronal synaptic networks due to changes in synaptic efficacy and circuit structure, and importance of EC L3 pyramidal neuron loss for these changes. This issues are studied using sharp electrode and patch clamp recording, Western Blot and fluorescent in-situ hybridization (FISH), and two-photon laser scan fluorescence microscopy of presynaptic vesicular release.